Our broad goal is to provide novel mechanistic insight into the pathogenesis of alcohol use disorders (AUDs) by ascertaining how Regulator of G protein signaling 6 (RGS6) functions in the mesolimbic circuit as a critical modulator of alcohol (EtOH) seeking and reward behaviors. EtOH is the most commonly abused drug worldwide and, despite decades of research, the neurobiological mechanisms underlying EtOH seeking behavior and dependence are not fully understood. As a result, there are few effective therapeutics to reduce alcohol cravings or withdrawal symptomology, and abstinence remains the only way to prevent AUDs. This proposal is based upon our discovery that RGS6 promotes mouse EtOH seeking and reward behaviors and on a subsequent GWAS identifying RGS6 as a gene linked to human AUDs. Based on our previous work, we hypothesized that RGS6 promotes EtOH seeking behavior by negatively regulating neuronal G?i/o-coupled receptors (GPCRs) implicated in alcoholism. Indeed, we found that RGS6-/- mice consume less EtOH when given free access and are less susceptible to EtOH reward, dependence and withdrawal. Antagonism of D2Rs or GABABRs and inhibition of dopamine transporter (DAT) activity, which is upregulated by G?i/o-coupled GPCRs, partially and completely restored the reduced EtOH consumption in RGS6-/- mice, respectively. Our findings suggest that by inhibiting GPCR-G?i-DAT upregulation in VTA DA neurons, RGS6 promotes EtOH-induced dopamine (DA) neurotransmission and behavioral responses. RGS6 control of this pathway would represent a previously unknown mechanism modulating the intensity and duration of G?i signaling in VTA neurons to titre DA levels responsible for acute and chronic behavioral effects of EtOH. Here we propose to test the central hypothesis that RGS6 regulation of G protein signaling in VTA DA neurons plays a critical role in promoting dopaminergic neurotransmission responsible for EtOH seeking and reward behaviors. Aim 1 will determine the role of mesolimbic RGS6 on EtOH seeking and reward behaviors using mice with selective deletion of RGS6 in VTA DA neurons as well as rescue studies with RGS6 or its G protein-defective mutant. Aim 2 will determine the role of RGS6 in VTA DA neurotransmission and on EtOH consumption in mice using optogenetics. We will employ state of the art cell type-specific optogenetics, neuronal ensemble recordings and Ca2+ imaging in the NAc, as well as a novel EtOH-triggered optogenetic approach to determine how RGS6 modulates mesolimbic DA neurotransmission and EtOH consumption in real time in freely moving mice. These studies are significant as they will illuminate an entirely novel pathway in the mesolimbic circuit, a major dopaminergic pathway in the brain implicated in EtOH behavioral reward and addiction. We will address a critical barrier to progress in the understanding and treatment of AUDs as RGS6 has been identified as a human AUD-linked gene and we seek to increase our basic knowledge of its role in the mesolimbic circuit.